Cancer Research Center, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, Hubei, P.R.China.
General Surgical Laboratory, The First Affiliated Hospital, Sun Yat-Sen University, Guangzhou, 510080, Guangdong, P.R.China.
Int J Biol Sci. 2021 Jul 5;17(11):2772-2794. doi: 10.7150/ijbs.60018. eCollection 2021.
Autophagy and glycolysis are two catabolic processes that manipulate pancreatic ductal adenocarcinoma (PDAC) development in response to hypoxia sensing, yet the underlying mechanism of how they are interlinked remain elusive. The functional roles of Unc-51 like kinase 1 and 2 (ULK1/2) in pyruvate kinase M2 (PKM2) transcription and glycolysis under hypoxia were assessed by chromatin immunoprecipitation, luciferase reporter, glucose consumption and lactate production assay. Co-immunoprecipitation, cellular ubiquitination, His-pulldown, protein kinase assay, immunofluorescence, immunohistochemistry, CRISPR technology, in silico studies were adopted to determine the molecular mechanism. Correlation analyses were performed in KPC (-Cre; LSL-Kras; Trp53) mice and clinical samples from PDAC patients. Therapeutic potential of ULK1/2 inhibitor and 2-deoxyglucose (2-DG) or 3-bromopyruvate (3-BP) was evaluated in cell-derived xenograft (CDX) and the patient-derived xenograft (PDX) models of nude mice. ULK1/2, but not ULK3, augments hypoxic glycolysis in PDAC cells mediated by PKM2 independent of BCL2/adenovirus E1B 19 kDa interacting protein 3 (BNIP3). Mechanistically, hypoxia stimulates ULK1 to translocate into nucleus, where it interacts with and phosphorylates yes-associated protein (YAP) at Ser227, resulting in YAP stabilization through blockade of ubiquitin-proteasome system (UPS), which in turn facilitates PKM2 transcription, glycolysis, cell proliferation as well as PDAC growth in mice. ULK1/2 is positively correlated with YAP and PKM2 in tumor tissues from KPC mice and clinical samples from PDAC patients. Pharmacological deactivation of ULK1/2 potentiates the antineoplastic efficacy of 2-DG and 3-BP in CDX and PDX models. Our findings underscore the Ser227 autophosphorylation-dependent nuclear YAP stabilization as a central node that couples ULK1/2-initiated autophagy to hypoxic glycolysis during PDAC development and propose that targeting ULK1/2 combined with 2-DG or 3-BP might be a feasible therapeutic strategy against PDAC.
自噬和糖酵解是两种分解代谢过程，它们通过感应缺氧来操纵胰腺导管腺癌 (PDAC) 的发展，但它们相互关联的潜在机制仍不清楚。通过染色质免疫沉淀、荧光素酶报告基因、葡萄糖消耗和乳酸产生测定评估了非典型卷曲相关激酶 1 和 2 (ULK1/2) 在缺氧下丙酮酸激酶 M2 (PKM2) 转录和糖酵解中的功能作用。采用免疫共沉淀、细胞泛素化、His 下拉、蛋白激酶测定、免疫荧光、免疫组织化学、CRISPR 技术、计算机模拟研究来确定分子机制。在 KPC(-Cre;LSL-Kras;Trp53) 小鼠和 PDAC 患者的临床样本中进行了相关性分析。在裸鼠细胞衍生异种移植 (CDX) 和患者衍生异种移植 (PDX) 模型中评估了 ULK1/2 抑制剂和 2-脱氧葡萄糖 (2-DG) 或 3-溴丙酮酸 (3-BP) 的治疗潜力。ULK1/2，但不是 ULK3，增强了 PDAC 细胞中由 PKM2 介导的缺氧糖酵解，而不依赖于 BCL2/腺病毒 E1B 19 kDa 相互作用蛋白 3 (BNIP3)。在机制上，缺氧刺激 ULK1 易位到核内，在核内与 yes 相关蛋白 (YAP) 相互作用并磷酸化 YAP 丝氨酸 227 位，通过阻断泛素-蛋白酶体系统 (UPS) 使 YAP 稳定，从而促进 PKM2 转录、糖酵解、细胞增殖以及小鼠体内的 PDAC 生长。ULK1/2 与 KPC 小鼠肿瘤组织和 PDAC 患者临床样本中的 YAP 和 PKM2 呈正相关。ULK1/2 的药理学失活增强了 2-DG 和 3-BP 在 CDX 和 PDX 模型中的抗肿瘤疗效。我们的研究结果强调了 Ser227 自身磷酸化依赖性核 YAP 稳定作为一个核心节点，将 ULK1/2 启动的自噬与 PDAC 发展过程中的缺氧糖酵解联系起来，并提出靶向 ULK1/2 联合 2-DG 或 3-BP 可能是对抗 PDAC 的一种可行的治疗策略。
